Reference
Complete the bibliographic reference for the article according to AJE format.

Humphries SE, Talmud PJ, Hawe E, Bolla M, Day INM, Miller GJ.  Apolipoprotein E4 and coronary heart disease in middle-aged men who smoke: a prospective study.  Lancet 2001;358:115-9.

Category of HuGE information
Specify the types of information (from the list below) available in the article:

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction
4. Gene-gene interaction
5. Genetic test evaluation/monitoring

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction

Hypothesis:  Smoking and APOE*ε4 are both associated with increased risk of coronary heart disease (CHD), and both have atherogenic effects on the oxidation of lipoprotein particles.  The association of smoking with CHD might therefore vary by APOE genotype.

Gene(s)
Identification of the following:

1. Gene name
2. Chromosome location
3. Gene product/function
4. Alleles
5. OMIM #

1. Gene name: APOE
2. Chromosome location: 19z13.2
3. Gene product/function: Apolipoprotein E, a principal apoprotein of chylomicrons, binds to receptors on liver cells and peripheral cells and facilitates uptake and catabolism of chylomicron fragments; also functions in protection of lipoproteins from oxidation
4. Alleles: ε2, ε3, ε4
5. OMIM#: 107741

Environmental factor(s)
Identification of the major environmental factors studied (infectious, chemical, physical, nutritional, and behavioral)

Smoking

Incident coronary heart disease (defined as fatal CHD events and nonfatal myocardial infarction [MI], based on WHO criteria), coronary artery surgery, and silent MI on follow-up ECG

1. Case-control
2. Cohort 
3. Cross-sectional
4. Descriptive or case series
5. Clinical trial
6. Population screening

Cohort

Cohort definition
For study designs 2, 3, and 6, the following are defined, where available:

1. Cohort selection criteria
2. Exclusion criteria
3. Gender
4. Race/ethnicity
5. Age
6. Time period
7. Geographic location
8. Number of participants

1. Cohort selection criteria: Healthy men aged 50-61 years registered in nine general medical practices in the UK
2. Exclusion criteria: “...free of history of unstable angina, myocardial infarction or evidence of silent infarction, coronary surgery, aspirin or anticoagulant therapy, cardiovascular disease, malignant disease (except skin cancer other than melanoma), or any condition precluding informed consent.”   Also excluded were nonwhites (not defined), subjects without DNA samples, and those with genotype ε2/ε4.
3. Gender: Male
4. Race/ethnicity: White
5. Age: 50-61 years at entry
6. Time period: Recruitment period not reported; follow-up duration not reported although reported person-years and sample size permit estimate of average duration = 8.3 years.
7. Geographic location:  United Kingdom (Camberly, Chesterfield, Halesworth, Harefield, North Mymms, Poole, St. Andrews)
8. Number of participants: 2,258

1. Environmental factor
2. Exposure assessment
3. Exposure definition
4. Number of participants with exposure data (% of total eligible)

1. Environmental factor: Smoking
2. Exposure assessment: Self-report
3. Exposure definition:
   a. Smokers: Smoking at least one cigarette daily on average for a year or more
   b. Ex-smokers: Smoking less than one cigarette a day
   c. Never smokers: Not defined
4. Number of participants with exposure data: N ( % of total eligible ) 234/236 patients ( 99% ) for cases and 634 ( 86% ) for controls.

1. Gene
2. DNA source
3. Methodology
4. Number of participants genotyped (% of total eligible) 

1. Gene: APOE
2. DNA source: Venous Blood
3. Methodology: “Salting out” with 6M NaCl and ethanol as described in Miller SA et al. Nucleic Acids Res 1988;16:1215.
4. Number of participants genotyped: 2303/3052 eligible (75%) but full data were available on 2258, after exclusion of 45 men with the ε2/ε3 genotype

1. Prevalence of gene variant
2. Gene-disease association
3. Gene-environment interaction
4. Gene-gene interaction
5. Genetic test evaluation/monitoring

1. Prevalence of APOE genotypes in subjects with triglyceride and smoking measures.
   

   Genotype	Number (%)
   ε3/ε3	1339 (60.5)
   ε2/ε2 and ε2/ε3	288 (13.0)
   ε3/ε3 and ε3/ε4	587 (26.5)

   
   
2. Gene-disease association: Not provided due to significant interaction with smoking
3. Gene-environment interaction:

Smoking is risk factor for CHD, particularly in men carrying the ε4 allele.  A gradient of increasing CHD risk from never smoker to ex-smoker to current smoker was observed in all genotype groups, but this gradient was greater in the ε4 group than in all others combined.

The study would have been stronger if CHD events had excluded coronary bypass surgery, though in the presumed period of follow-up surgical referrals might not have been expected to be strongly influenced by knowledge of APOE genotype.

Completeness of follow-up was not reported.